We developed a convenient and inexpensive method for the collection of buccal cells as a source of DNA in studies of school aged children. The method yields a large amount of high quality DNA. We used the method in our studies in Wuhan China and Southern California. We published a paper on this method in the past year. In the Wuhan study, we found that children living in homes were coal was burned for cooking or heating have an increased prevalence of wheezing. Children living with smokers are at increased risk of cough and phlegm production. This Chinese population is of special interest for the study of health effects of environmental tobacco smoke because there are very few mothers who smoke. Most of the exposure comes from smoking fathers. We have also found that mold exposure is the strongest risk factor in this study of wheezing, cough and phlegm. We are also beginning a collaboration with Dr. Darryl Zeldin to look at determinants of Alternaria exposure and the relation between Alternaria levels and asthma in the National Allergen Survey, an NIEHS study of home allergens. This work is part of the thesis work, along with the Wuhan study, for a UNC environmental health sciences study, Paivi Salo, an NIEHS predoctoral IRTA. The aim of this project is to examine genetic and environmental factors that influence the risk of childhood respiratory illnesses and to evaluate gene-environment interactions. To this end, we have assembled a set of three international studies of school aged children from populations with three different asthma prevalences: Wuhan China (low), Mexico City (intermediate) and Southern California, US (high). The three sites provide a range of exposures to environmental factors that are likely to interact with genetic susceptibility. These include ozone, endotoxin and environmental tobacco smoke. Notable exposures in the Wuhan China population are indoor coal burning, high prevalence of environmental tobacco smoke, and ambient air pollution. The Mexico City site is notable for the highest levels of ozone in North America and endotoxin levels are also high. The Southern California study offers a range of well characterized exposures to air pollution. The three studies in this project are conducted among school aged children with slightly different designs. The Wuhan study is a school-based study of 5,051 seventh graders at 22 schools enrolled in 1999. We collected environmental exposure and respiratory outcome data, pulmonary function and DNA from buccal cells. We are currently following the students for the third year. The Mexico City study uses the case-parent triad design. We are enrolling asthmatic children aged 7-17 at an inner city hospital along with their parents as controls. The case-parent triad design enables the examination of candidate gene associations from bias due to ethnic stratification, a potential problem in the two population mix of Mexico City. The case-parent triad design also allows the discernment of fetal versus maternal genetic effects. This Mexico City population experiences the highest ozone levels in North America. Ambient endotoxin is also elevated. We are examining candidate genes involved in respiratory responses to ozone, based on human and animal evidence. The Southern California study is a school-based air pollution cohort study that I helped establish before coming to NIEHS. Beginning in the spring of 1998, I initiated a collection of genetic material on the cohort to examine candidate genes for asthma and impaired growth of pulmonary function. This collection was so successful that it was incorporated into an NIEHS/EPA Children's Center grant and is now extramurally funded. In the Southern California study, out main finding this year was that children with the highest levels of outdoor exercise were at increased risk of developing asthma if they lived in high ozone communities. This is one of the first prospective studies to suggest that ozone exposure may be a risk factor for asthma. We also found ozone exposure was related to higher levels of school absence. In another type of analysis we found that children relocating from communities with lower PM10 to those with higher PM10 experienced decreased growth in lung function. Together these studies help to establish adverse effects of air pollution in growing children. We have established an effective and efficient means of collecting a large amount of high quality genetic material on children in school based studies. This published work should be useful to investigators planning similar studies. We are continuing subject enrollment in our case-parent traid study of asthma in Mexico City. To date, we have enrolled over 250 complete triads. DNA extraction is complete on these samples and we have examined samples for polymorphisms in genes that are plausibly involved in response to ozone. We have selected genes based on results from mouse models of ozone effects as well as mechanistic studies of ozone effects. Genes include IL-13, IL-6, IL-8, tnf-alpha, GSTP1, NQ01 and myeloperoxidase. We are continuing with subject enrollment and genetic analyses and hope to complete data collection by early FY 2004. In preliminary analyses on IL-13, an increased risk was seen for two linked promoter variants. The results are in the direction predicted based on functional effects of these polymorphisms. In the Wuhan study, we have identified several risk factors for respiratory illness among Chinese adolescents -- enviromental tobacco smoke, indoor coal burning and exposure to molds. There are few data on risk factors for respiratory illness in this population. The population is interesting because most of the environmnental tobacco smoke exposure is via the fathers. Few mothers smoke. Nonetheless, health effects of environmental tobacco smoke were clearly seen. This population is also of interest because coal burning remains a common exposure.